This proposal represents an attempt to learn the biochemical basis of two regulatory phenomena, S1 and S2, controlled by bacteriophage T4. Upon infection of E. coli by wild-type T4, the synthesis of a large number of "early" enzymes (involved in DNA metabolism) proceeds until about 12 min at 37 degrees and then shuts off; we term this shutoff S1. If infection is carried out with a mutant unable to make "late" (maturation) proteins, S1 shutoff fails and a second shutoff (S2) activates at 20 min; such mutants include a large number unable to make DNA, and those defective in genes 33 and 55. We have discovered a mutant, SP62, which specifically lacks the S2 shutoff for certain early enzymes; this mutant defines a new T4 gene, regA. Several lines of evidence argue that the regA gene exerts its control after transcription; we favor the notion that it affects the specificity of translation of mRNA. Evidence exists that the S1 control is also post- transcriptional. In Project I we will compare ribosomes (and their associated initiation factors) isolated after infection by wild T4, and mutants in genes regA, 55, 33 and several DNA-synthetic genes; T4- specific proteins in the ribosomes will be radioactively labeled and separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and detected in the slab gels by autoradiography. We will attempt to identify any proteins that differ among the patterns. In Project II, we will pursue further our promising preliminary indications that the regA gene is at least partly responsible for the failure of wild-type, interparental recombinant DNA to be expressed, in infections of a non-permissive host by two different DNA- amber mutants in the same early gene. In Project III, SDS-PAGE will be used to study why the regA mutant is defective in maturation of phage DNA at high temperature. In Project IV, the regA mutant serves as a tool in a scheme to select mutants specifically defective in the S1 shutoff. If found, they will be studied in much the same way we are studying the regA mutant.